Advances in microfluidics technology and high performance liquid chromatography are revolutionizing molecular biology procedures for enzymatic analysis (e.g., glucose and lactate assays), DNA analysis (e.g., polymerase chain reaction and high-throughput sequencing), and proteomics. The basic idea of microfluidic biochips is to integrate assay operations such as detection, sample pre-treatment and sample preparation on a single microchip. An emerging application area for biochips is clinical pathology, especially the immediate point-of-care diagnosis of diseases. In addition, microfluidics-based devices, capable of continuous sampling and real-time testing of air/water samples for biochemical toxins and other dangerous pathogens, can serve as an always-on “bio-smoke alarm” for early warning. Low flow separation techniques, such as capillary electrophoresis, capillary electrochromatography, and low flow HPLC & UHPLC are further emerging applications.
A lab-on-a-chip (LOC) is a device that integrates one or several laboratory functions on a single chip from a few millimeters to a few square centimeters in size. LOCs deal with the handling of extremely small fluid volumes down to less than pico liters. Lab-on-a-chip devices are a subset of Microelectromechanical Systems (MEMS) devices and are often indicated by “Micro Total Analysis Systems” (μTAS) as well. Microfluidics is a broad term that includes mechanical flow control devices like pumps, valves and sensors such as flow meters and viscometers. “Lab-on-a-Chip” generally relates to the scaling of single or multiple lab processes down to chip-format, whereas “μTAS” is dedicated to the integration of the total sequence of lab processes to perform chemical analysis.
μTAS technologies are suitable for applications other than just analysis. For example, channels (capillary connections), mixers, valves, pumps and dosing devices are all suitable μTAS technologies.
The first LOC analysis system was a gas chromatograph, developed in 1975 by S. C. Terry—Stanford University. However, it was not until the end of the 1980's, and beginning of the 1990's, that LOC research started to seriously grow. The development of micropumps, flow sensors and the concepts for integrated fluid treatments for analysis systems was spurred by this research. These μTAS concepts demonstrated that integration of pre-treatment steps, usually done at lab-scale, could extend the simple sensor functionality towards a complete laboratory analysis, including additional cleaning and separation steps.
A big boost in research and commercial interest came in the mid 1990's, when μTAS technologies turned out to provide interesting tooling for genomics applications such as capillary electrophoresis and DNA microarrays. Another boost in research support came from the military, especially from DARPA (Defense Advanced Research Projects Agency), for their interest in portable bio/chemical warfare agent detection systems. The added value was not only limited to integration of lab processes for analysis but also the characteristic possibilities of individual components and the application to other, non-analysis, lab processes. Hence the term “Lab-on-a-Chip” was introduced.
Although the application of LOCs is still novel and modest, a growing interest of companies and applied research groups is observed in different fields such as analysis (e.g. chemical analysis, environmental monitoring, medical diagnostics and cellomics) but also in synthetic chemistry (e.g. rapid screening and microreactors for pharmaceutics). Further application developments, research in LOC systems is expected to extend towards downscaling of fluid handling structures as well, by using nanotechnology. Sub-micrometer and nano-sized channels, DNA labyrinths, single cell detection analysis and nano-sensors are feasible for interaction with biological species and large molecules.
Despite the immense amount of research around creating the chips themselves, interfacing to the real world, the “Chip-to-World” interface technology, has been limited. Progress to interface to the LOCs has progressed slowly. This invention serves as a way to make connections to microchips and similar-based microfluidic devices.
Lab-on-a-chip technology may be used to improve global health, particularly through the development of point-of-care testing devices. In countries with few healthcare resources, infectious diseases that would be treatable in developed nations are often deadly. In some cases, poor healthcare clinics have the drugs to treat a certain illness but lack the diagnostic tools to identify patients who should receive the drugs. LOC technology may be the key to provide powerful new diagnostic instruments. The goal of these researchers is to create microfluidic chips that will allow healthcare providers in poorly equipped clinics to perform diagnostic tests such as immunoassays and nucleic acid assays without additional laboratory support.
The basis for most LOC fabrication processes is photolithography. Initially most processes were in silicon, as these well-developed technologies were directly derived from semiconductor fabrication. Because of demands, for, e.g., specific optical characteristics, bio- or chemical compatibility, lower production costs and faster prototyping, new processes have been developed such as glass, ceramics and metal etching, deposition and bonding, PDMS processing (e.g., soft lithography), thick-film- and stereolithography as well as fast replication methods via electroplating, injection molding and embossing. Furthermore, the LOC field more and more exceeds the borders between lithography-based microsystem technology, nanotechnology and precision engineering.
LOCs may provide advantages, which are specific to their application. Typical advantages of LOC systems include:
low fluid volumes consumption (less waste, lower reagents costs and less required sample volumes for diagnostics);
faster analysis and response times due to short diffusion distances, fast heating, high surface to volume ratios, small heat capacities;
better process control because of a faster response of the system (e.g. thermal control for exothermic chemical reactions);
compactness of the systems due to integration of much functionality and small volumes;
massive parallelization due to compactness, which allows high-throughput analysis;
lower fabrication costs, allowing cost-effective disposable chips, fabricated in mass production; and
safer platform for chemical, radioactive or biological studies because of integration of functionality, smaller fluid volumes and stored energies.
To interface microchips to their supporting hardware systems remains a significant challenge. And the lack of robust, reliable technology to make these connections has not only slowed microfluidic research, but is preventing chip-based solutions from being applied to real world applications. While extensive research effort has been directed toward microchip performance and fabrication, very little effort has been focused on technologies to interface these chips to fluidic and electronic hardware. The end result is that microchip performance is often compromised due to the underdeveloped interface technology.
Nano-liquid chromatography (nanoLC) is also a powerful technique that has significant challenges. NanoLC uses chromatography columns with inner diameters ranging from 25-150 μm packed with 2-5 μm stationary phase particles. However, the most typical column size is 75 μm inner diameter with <5 μm particles. Typical nanoLC flow rates range from 50-300 nL/min. Smaller particle sizes and longer columns generate higher resolving power, but also increased backpressure on the system. NanoLC is ideal for resolving highly complex, intractable biological mixtures. This is due to the fact that nanoLC only requires attomole to femtomole sample amounts and offers high sensitivity because of its resolving power. As a result, more complex problems may now be addressed such as molecular interactions, ion structures, quantitation, and kinetics in the both the field of proteomics and glycomics. Consequently, nanoLC is a necessity for biological laboratories. Other low flow separation techniques including capillary electrophoresis, capillary zone electrophoresis, and capillary electrochromatography offer high sensitivity, but are difficult to couple to mass spectrometry and have limited sample loading volumes.
The combined technique of nanoLC/electrospray/mass spectrometry, often abbreviated nanoLC/MS, has emerged as the gold standard for proteomic and glycomic laboratories. This combined technique can resolve highly complex mixtures with components covering a wide dynamic range, can then obtain valuable mass spectral data, and ultimately identify the components in the mixture. Furthermore the technique can identify, localize, and structurally characterize subtle chemical variations between sample components such as post-translational modifications. Quantitative proteomic profiling using LC/MS is an emerging technology with great potential for the functional analysis of biological systems and for the detection of clinical diagnostic marker proteins. This technology has been demonstrated for quantitation of proteins, as well as specifically for phosphoproteins and glycoproteins. In addition to protein identification, characterization of post-translational modifications, and quantitation of protein differential expression, nanoLC/MS has also been used to investigate protein-protein complexes. Thus, nanoLC/MS is a far-reaching technology, positively impacting many areas of proteomics, and consequently the technique is invaluable to biological laboratories. However, unfortunately nanoLC/MS is alarmingly underutilized due to the complexity, limited robustness, and high level of expertise required of nanoLC/ESI systems. These unfavorable attributes arise from several shortcomings of the technique.
Conventional nanoLC/ESI systems suffer from several limitations. The first drawback is that system reproducibly is highly dependent on user skill level. This is due to user inabilities handling, cutting, connecting, positioning, and inconsistently setting-up the column and spray emitter. The second limitation is the level of difficulty in making the required nanofluidic connections. The delicate nanoLC connections are dependent on individual human coordination skills. In addition, conventional fittings frequently fatigue, slip or loosen over time and require further tightening, but all too often fittings are over-tightened which leads to leakage. Also, hand-cutting of capillaries often leads to formation of fractures and jagged ends of the cleaved tubing, which can produce thousands of fused-silica particulates when connections are attempted. These particulates cause column and emitter clogging and plugging, as well as internal valve damage. For low-pressure, post-column connections, often press-tight connectors are used. These press-tight connectors are notorious for leaking, and for plugging capillaries as teflon, from the teflon tubing, is shaved from the interior connector walls when the capillary is inserted. A poor connection can also create dead volume, which reduces chromatographic resolution. Then to troubleshoot the system when there is poor performance or a malfunction is very difficult. For small fluidic leaks, visual assessment and diagnosis is often not possible. High user skill is needed to troubleshoot, and repairing the system frequently involves blind substitution of parts to diagnose the problematic components. Finally, the technique is very labor intensive, especially when the system is first started.
So called “zero-dead-volume” couplings attempt to minimize the amount of unswept area at the coupling. Unfortunately, “zero-dead-volume” fittings still allow the formation of voids and unswept volumes in the area where the tube and the sealing feature of the fitting meet. Further, zero-dead-volume fittings are difficult to manufacture and, in the case of a chromatograph, allow exposure of the material coating the tube that absorbs and retains components of the chromatographic sample flow. Therefore, it would be desirable to provide a fluidic coupling from a tube to a fluidic path. Additionally the sealing connection may be into a component other than a cylindrical bore, such as many microfluidic devices.
In summary, microfluidics and nano-liquid chromatography, are powerful techniques with tremendous challenges. The majority of these challenges reside in the making of leak-tight fluidic connections. The invention disclosed here provides a solution to facilitate the making of leak-tight fluidic connections for applications including microfluidics and liquid chromatography.